2023, 31(4):329-335, 342.DOI: 10.20039/j.cnki.10073949.2023.04.007
Abstract:Aim To investigate the effect of Crocin on hypoxia/reoxygenation injury of cortical neurons and its molecular mechanism. Methods The cortical neurons of SD rats were isolated and cultured, and the control group, model group, Crocin (50 mg/L) group, Crocin (50 mg/L)+lipopolysaccharide (TLR4 activator, 100 μg/L) group were set. The cortical neurons in control group were routinely cultured; the cortical neurons in model group were given hypoxia 4 h and reoxygenation 24 h; the cortical neurons in Crocin group and Crocin+lipopolysaccharide group were intervened for 24 h, and then the model was established. The neuronal viability and apoptosis rate were detected by CCK-8 method and Annexin V-FITC/PI double staining method. The levels of inflammatory factors in the culture medium were detected by ELISA method. The expression of toll-like receptor 4 (TLR4), nuclear factor-κB p65 (NF-κB p65), p-NF-κB p65, inhibitor α of NF-κB (IκBα), p-IκBα, high mobility group box protein B1 (HMGB1), cleaved Caspase-3,Bü lymphoma-2 gene (Bcl-2), Bcl-2-associated X protein (Bax) were detected by Western blot method. Results Compared with the control group, the activity of cortical neurons in the model group decreased significantly (P<0.05); the apoptosis rate, the levels of tumor necrosis factor-α (TNF-α), interleukin-1β (IL-1β), IL-6 in the culture medium, the expressions of TLR4, HMGB1, cleaved Caspase-3 and the ratio of p-NF-κB p65/NF-κB p65, p-IκBα/IκBα, Bax/Bcl-2 increased significantly (P<0.05). Compared with the model group, the activity of cortical neurons in Crocin group increased by 94.97% (P<0.05); the apoptosis rate decreased by 65.80% (P<0.05); the levels of TNF-α, IL-1β, IL-6 in the culture medium decreased by 61.86%, 78.34%, 63.42% (P<0.05); the expressions of TLR4, HMGB1, cleaved Caspase-3 and the ratio of p-NF-κB p65/NF-κB p65, p-IκBα/IκBα, Bax/Bcl-2 decreased by 73.43%, 52.13%, 81.52%, 69.70%, 60.55%, 95.05% (P<0.05). Lipopolysaccharide significantly reversed the regulatory effects of Crocin on hypoxia/reoxygenation injured cortical neurons (P<0.05). Conclusion Crocin can inhibit the apoptosis and inflammatory response of hypoxia/reoxygenation injured neurons, and has protective effect on hypoxia/reoxygenation injury of cortical neurons, which mechanism may be related to the inhibition of TLR4/NF-κB signaling pathway activation.
2021, 29(5):383-388.
Abstract:Aim To investigate the protective effect of Crocin on myocardial mitochondria in rats with acute myocardial infarction (AMI) and its mechanism. Methods 150 SD rats were randomly divided into Sham group, AMI group, Crocin (7.5,5, 30 mg/kg) group according to random number table method (n=30). The AMI rat model was established by ligating the left anterior descending coronary artery. 24 h later, the ultrastructure changes of myocardial mitochondria in ischemic area was observed by transmission electron microscope; the membrane potential and the opening of mitochondrial permeablity transition pore (MPTP) were detected by fluorescence spectrophotometer; the myocardial mitochondrial respiratory function index (R3, R4, RCR) were detected by oxygen electrode method. The respiratory enzymes activity, ATP content were detected by colorimetry method; the Na+-K+-ATPase, Ca2+-ATPase activity were detected by phosphorus determination method; the mitochondrial Ca2+ concent was detected by atomic chemiluminescence. ResultsCompared with AMI group, the ultrastructural lesions such as mitochondrial swelling, membrane rupture, crest rupture and dissolution were significantly improved in Crocin 5,0 mg/kg groups, the membrane potential increased and MPTP openness decreased (P<0.01); the R3, RCR increased and R4 decreased (P<0.05 or P<0.01); respiratory enzymes (NADH dehydrogenase, succinate dehydrogenase, cytochrome C oxidase) activity, ATP content and Na+-K+- ATPase, Ca2+-ATPase activity increased (P<0.05 or P<0.01), Ca2+ concentration decreased (P<0.05 or P<0.01). Conclusion Crocin has protective effect on the structure and function of myocardial mitochondria in the ischemic area of AMI rats, which mechanism may be related to inhibiting the increase of mitochondrial Ca2+ concentration.